The ultimate goal of this application is to identify the host signaling pathways targeted by the type III secretion system 1 (T3SS1) of the gram-negative bacterium Vibrio parahaemolyticus; a major agent responsible for gastroenteritis outbreaks associated with the consumption of contaminated seafood. The prevalence of V. parahaemolyticus in the environment and incidence of infection have been linked to rising water temperatures caused by global warming. A T3SS is a needle-like structure that efficiently translocates virulence factors from bacteria into the cytosol of a host cell. The virulence factors (also called bacterial effectors) have evolved in a manner similar to many of the viral oncogenes; a eukaryotic activity is usurped and modified by the pathogen for its own advantage. Genomic sequencing of V. parahaemolyticus revealed the existence of two pathogenicity islands that encode both a T3SS and putative effectors. The first pathogenicity island contains T3SS1 and is associated with a cytotoxicity, while the second pathogenicity island contains T3SS2 and is associated with an enterotoxicity. The effectors on the first pathogenicity island have been identified as open reading frames with no obvious homology to any known protein. Recently, we have demonstrated that the V. parahaemolyticus uses T3SS1 to orchestrate a multifaceted host cell infection by induction of autophagy, cell rounding and then cell lysis. We have shown one of the T3SS1 effectors, VopS, utilizes a novel posttranslational modification called AMPylation to disrupt host signaling. Herein, we propose Specific Aims to study the multifaceted death induced by T3SS1 by identifying the activity and host targets of three known effectors, identify other effectors secreted by T3SS1 and characterize the host signaling pathways that are disrupted by V. parahaemolyticus during infection. These studies provide molecular insight into the molecular mechanisms used by this harmful gastrointestinal bacterial pathogen V. parahaemolyticus.